Department of Pathology, Stanford University School of ......DNA and RNA in respiratory secretions (IDbyDNA, San Francisco, CA) [1]. Of these, the Karius test has been commercially

© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society

of America. All rights reserved. For permissions, e-mail: [email protected].

Clinical Impact of Metagenomic Next-Generation Sequencing of Plasma Cell-Free DNA

for the Diagnosis of Infectious Diseases: A Multicenter Retrospective Cohort Study

Catherine A. Hogan1,2,3, Shangxin Yang4, Omai B. Garner4, Daniel A. Green5, Carlos A.

Gomez6, Jennifer Dien Bard7, Benjamin A. Pinsky1,2,3,8, Niaz Banaei1,2,3,8

1 Department of Pathology, Stanford University School of Medicine, Stanford, CA, USA

2 Clinical Microbiology Laboratory, Stanford Health Care, Stanford, CA, USA

3 Clinical Virology Laboratory, Stanford Health Care, Stanford, CA, USA

4 Department of Pathology and Laboratory Medicine, University of California Los

Angeles, Los Angeles, CA, USA

5 Department of Pathology, Columbia University Irving Medical Center, New York, NY,

USA

6 Division of Infectious Diseases, Department of Medicine, University of Utah, Salt Lake

City, UT, USA

7 Department of Pathology, Children’s Hospital of Los Angeles, Los Angeles, USA

8 Division of Infectious Diseases and Geographic Medicine, Department of Medicine,

Stanford University School of Medicine, Stanford, CA, USA

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ic.oup.com/cid/advance-article-abstract/doi/10.1093/cid/ciaa035/5703656 by Florida H

ospital Medical Library user on 15 January 2020

2

Corresponding author:

Niaz Banaei

3375 Hillview, Room 1602

Palo Alto, CA 94304

Phone (650) 736-8052

Fax (650) 725-5671

[email protected]

Summary: In a multicenter retrospective cohort study, we show that the real-world

clinical impact of plasma metagenomic next-generation sequencing (mNGS) for the

noninvasive diagnosis of infections is limited (positive impact 7.3%). Further studies are

needed to optimize the impact of mNGS.

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Abstract

Background: Metagenomic next-generation sequencing (mNGS) of plasma cell-free

DNA has emerged as an attractive diagnostic modality allowing broad-range pathogen

detection, noninvasive sampling, and earlier diagnosis. However, little is known about

its real-world clinical impact as used in routine practice.

Methods: We performed a retrospective cohort study of all patients for whom plasma

mNGS (Karius test) was performed for all indications at 5 U.S. institutions over 1.5

years. Comprehensive chart review was performed, and standardized assessment of

clinical impact of the mNGS based on the treating team’s interpretation of Karius results

and patient management was established.

Results: A total of 82 Karius tests were evaluated, from 39 (47.6%) adults and 43

(52.4%) children and a total of 53 (64.6%) immunocompromised patients. Karius

positivity rate was 50/82 (61.0%), with 25 (50.0%) showing two or more organisms

(range, 2-8). The Karius test results led to positive impact in 6 (7.3%), negative impact

in 3 (3.7%), no impact in 71 (86.6%), and was indeterminate in 2 (2.4%). Cases with

positive Karius result and clinical impact involved bacteria and/or fungi but not DNA

viruses or parasites. In 10 patients who underwent 16 additional repeated tests, only

one was associated with clinical impact.

Conclusions: The real-world impact of the Karius test as currently used in routine

clinical practice is limited. Further studies are needed to identify high-yield patient

populations, define the complementary role of mNGS to conventional microbiological

methods and how best to integrate mNGS into current testing algorithms.

Key words: metagenomic next-generation sequencing, plasma

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Introduction

Metagenomic next-generation sequencing (mNGS) of pathogen nucleic acid in

clinical samples has emerged as a promising one-test approach for hypothesis-free

diagnosis of potentially all infectious etiologies. mNGS is currently orderable in select

reference laboratories from plasma cell-free DNA (Karius, Redwood City, CA), from

DNA and RNA in cerebrospinal fluid (CSF) (University of California, San Francisco) and

DNA and RNA in respiratory secretions (IDbyDNA, San Francisco, CA) [1]. Of these,

the Karius test has been commercially available the longest and is likely the most

common mNGS send-out test in U.S. institutions. This assay is reported to detect and

quantify pathogen cell-free DNA from 1,250 bacteria, DNA viruses, fungi and eukaryotic

parasites [2], and has been employed for the diagnosis of invasive fungal disease [3, 4],

community-acquired pneumonia [5] and infections in immunocompromised hosts [6, 7].

The availability of clinical metagenomic cell-free DNA sequencing has generated

interest from clinical providers across disciplines given the noninvasive nature of testing,

and the potential to pose an actionable diagnosis faster than through conventional

microbiologic methods. However, little is known about the clinical impact of mNGS

testing when used for routine patient care. A study assessing the clinical impact of

mNGS from CSF in patients with suspected meningitis and encephalitis showed positive

clinical impact in 8/204 (3.9%) patients tested [8]. In the present multicenter study, our

aim was to comprehensively examine the real-world clinical impact of the Karius test

ordered for all indications for the diagnosis of infectious diseases.

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Methods

Ethics. This study was approved by the Institutional Review Boards (IRB) at Stanford

University and Columbia Medical Center (CUMC). IRB was waived for this project for

University of California Los Angeles (UCLA), University of Utah (UT) and Children’s

Hospital of Los Angeles (CHLA).

Karius Test. The Karius laboratory started to offer commercial testing in December

2016. Plasma from whole blood collected in a K2-EDTA or BD Vacutainer PPT tube

(Becton Dickinson, Franklin Lakes, NJ) and separated <6 hours after draw was sent

either fresh (within 4 days of blood draw) or frozen. All testing was performed by Karius

per routine testing protocol as previously described [2].

Study Design. We performed a retrospective cohort study of patients for all

consecutive plasma mNGS samples sent-out for Karius test from 5 U.S. institutions

(CHLA, CUMC, Stanford Health Care, UCLA and UT) from August 1st 2017 to May 31st

2019. Comprehensive chart review was performed by each site investigator for

respective patients, and clinical impact solely based on the treating team’s interpretation

of Karius results and their subsequent management decisions was assessed according

to predefined objective grading criteria (Table 1) as to whether Karius test result had a

positive, negative, or no clinical impact, or if impact was indeterminate. This study

intended to assess the real-world impact of Karius test and therefore only the treating

team’s decisions and actions were considered for impact assessment, and no

retrospective clinical consensus assessment of impact and adjudication by a panel of

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experts was performed. Given that Karius testing may be reordered in the same patient

for different reasons including patient monitoring, the main analysis was performed on

the first Karius test per patient. A separate analysis was performed on repeat tests.

Statistical Analysis. Statistical analysis was performed with Stata 15 (Stata Corp, TX,

USA). Due to the descriptive nature of the review, a formal sample size calculation was

not performed.

Results

A total of 82 Karius tests were performed in unique patients cared for at UCLA

(n=50 tests), SHC (n=21), CUMC (n=5), UT (n=3) and CHLA (n=3) (Table 2). There

were 39 (47.6%) adults and 43 (52.4%) children, 52 (63.4%) males and 53 (64.6%)

immunocompromised patients, most commonly solid organ transplant (17.1%) and

primary immunodeficiency (13.4%) (Table 2). The mean turn-around-time from sample

collection to result was 3.1 days (SD: 1.2), and time from Karius receipt of sample to

result was 1.5 days (SD: 0.9) for a subset of 20 samples. The most common ordering

providers were infectious disease physicians (74.4%), followed by oncologists (11.0%).

The main indication for testing was fever of unknown origin (23.2%), suspected

respiratory infection (13.4%), sepsis (9.8%), suspected endocarditis (8.5%) and febrile

neutropenia (7.3%). The Karius positivity rate was 50/82 (61.0%), with 25 (50.0%) of

these tests resulted as monomicrobial (17 bacterial, 5 viral and 3 fungal), and 25

(50.0%) showing 2 or more organisms (range, 2-8). There was no parasite detected in

this cohort.

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The Karius test results led to positive or negative clinical impact in 9 (11.0%)

patients (Table 3). The impact was positive in 6 (7.3%) and negative in 3 (3.7%) (Table

3). Of these 9 Karius tests, 8 had organisms detected while 1 had no organism

detected. Impact was none in 71 (86.6%) tests and indeterminate in 2 (2.4%). All 8

cases with positive Karius result and clinical impact involved bacteria and/or fungi;

determination of the presence or absence of DNA viruses by plasma mNGS

demonstrated no clinical impact in this study. Patients with a positive or negative Karius

result where there was no (n=71) or indeterminate (n=2) clinical impact are presented in

Supplementary Tables 1 and 2. Positive clinical impact was categorized as de-

escalation of therapy (n=1), earlier diagnosis than conventional methods and initiation of

appropriate therapy (n=2), new diagnosis not made by conventional methods and

escalation of therapy (n=1), and new diagnosis and de-escalation (n=2) (Table 3). The 3

new diagnoses made by Karius test included presumptive diagnosis of an abdominal

aorta mycotic aneurysm caused by Enterococcus faecalis and Prevotella

melaninogenica, a diagnosis of Rhizopus oryzae confirming disseminated

mucormycosis, and identification of mixed oral flora in a case of blood culture-negative

endocarditis, facilitating de-escalation of therapy. The 3 cases associated with negative

clinical impact resulted in additional, unnecessary diagnostic investigations (n=2), and

unnecessary treatment (n=1). Among positive Karius test results, lack of clinical impact

was most commonly due to identification of a new organism that was not acted upon

(n=20), confirmation of conventional microbiological result that was not acted upon

(n=14), or both (n=6) (Supplementary Table 1).

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All patients in this cohort underwent both conventional testing and Karius test

send-out in the order decided at the discretion of the treating team. A total of 27/82

(32.9%) patients underwent Karius testing with a pre-established microbiological

diagnosis through conventional testing, with relative proportions varying by institution

from 0 to 60%. The most common conventional diagnostic method establishing a

microbiological diagnosis in this group was blood culture (n=9), tissue bacterial culture

(n=7) and viral blood-based PCR (n=4). Of the 27 patients with a positive conventional

diagnosis, 16 (59.3%) showed the same organism detected by the Karius test, with 7 of

these showing other organisms in addition to the one that was reproducibly detected

(Supplementary Tables 1 and 2). The remaining 11 cases where preceding

conventional microbiological testing was positive but Karius testing was negative

included bacteria (Staphylococcus aureus (n=1), Streptococcus viridans (n=2),

Mycobacterium haemophilum (n=1), Mycobacterium chelonae (n=1), Mycobacterium

bovis (n=1)), DNA virus (adenovirus (n=1)), and fungi (Candida parapsilosis (n=1),

Candida lusitaniae (n=1), Coccidioides immitis/posadasii (n=1), Mucorales (n=1)).

A subset of 10 patients underwent repeat testing, for a total of 16 additional

repeated tests from UCLA (n=14), SHC (n=1) and CUMC (n=1). Review of the 16

repeat tests revealed 15 positive cases of which 1 case was associated with a positive

clinical impact, where the repeat Karius test was performed 3 months after the initial

one. This 2-year-old boy with a history of heart transplant presented with positive blood

and respiratory cultures for Stenotrophomonas maltophilia. Four days after these

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conventional tests were performed, Karius test was sent-out and resulted two days later

as positive for Stenotrophomonas maltophilia. The absence of any other detected

organism on the Karius test prompted the treating team to de-escalate therapy.

Discussion

Metagenomic next-generation sequencing has stimulated much interest among

providers across medical subspecialties given its potential for broad-range pathogen

detection. Furthermore, the noninvasive nature of plasma samples makes mNGS an

attractive diagnostic modality for deep-seated infections that currently require invasive

biopsy. However, the value of mNGS testing as a diagnostic tool is poorly understood.

In this multicenter retrospective cohort study, we show that, as currently used in clinical

practice, the real-world clinical impact of the Karius test is limited given that it resulted in

no or negative impact in the majority (92.7%) of patients. This finding was consistent

with that reported for mNGS on CSF where 196 of 204 (96.1%) tests did not have a

clinical impact [8]. As it currently stands, in most academic and private health systems,

microbiology and virology diagnostic laboratories offer a comprehensive menu of

culture-based and molecular tests such that only few, typically complex, cases with

infectious etiology will be left without a diagnosis. Important questions to consider now

are whether mNGS has the diagnostic accuracy required to complement currently-

available diagnostics, and how best to integrate mNGS into current testing algorithms.

Published studies indicate CSF and plasma mNGS assays lack sensitivity compared to

conventional methods and therefore cannot be used as stand-alone or rule-out tests [2,

8]. Thus, whether reserving mNGS as last resort for cases for which conventional

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microbiological testing has failed to provide an answer, or to include mNGS more

proximally in testing in parallel to conventional tests is currently an active subject of

discussion in the microbiology field that needs to be determined empirically [9]. The

decision to include mNGS testing should take into account the differential diagnosis, as

well as the availability, accuracy, and turnaround time of conventional methods as well

as cost-effectiveness of mNGS. The specific example of lack of clinical impact of

positive Karius results for DNA viruses in this cohort provides a compelling case for

routine highly-available, highly-accurate molecular virology assays providing sufficient

and actionable diagnostic information in all cases.

Plasma cell-free DNA may originate from the site of infection or colonization.

Although Karius reports quantitative units, there is no threshold that distinguishes

colonization from infection, and interpretation of results may therefore be clinically

challenging, especially in immunocompromised hosts and patients with disrupted

intestinal barrier. Clinical validation data recently reported on the Karius test from

patients presenting to the emergency department at a tertiary hospital with suspected

sepsis have demonstrated a positive percent agreement of 84.8% and negative percent

agreement of 48.2% compared to all microbiological reference testing [2]. The finding

that only 6 of 50 (12.0%) positive test results were positively impactful in the current

study is consistent with an assay specificity that is insufficient to enable clinical impact

when organisms are detected under routine practice. Even in certain cases where care

was tailored to the positive Karius result, it is unclear if the reported organisms were the

underlying drivers of pathology. For example, the patient with abdominal aorta mycotic

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aneurysm (Table 3; Study ID SHC-11) in whom antibiotic therapy was de-escalated to

chronic suppressive therapy with amoxicillin-clavulanic acid based on a positive Karius

result with Enterococcus faecalis and Prevotella melaninogenica presented with multiple

clinical recurrences of back pain and eventually grew Mycobacterium avium complex

(MAC) from a vertebral tissue biopsy 9 months later.

Patient selection practices may also have contributed to low clinical impact as

almost one-third (ranging from 0 to 60% by institution) of patients had already received

a microbiological diagnosis prior to testing. The only case in this cohort to have

demonstrated clinical impact in the setting of a pre-established microbiological

diagnosis is that of the repeat Karius test identifying Stenotrophomonas maltophilia only

in a patient with known bacteremia with the same organism, leading to de-escalation of

therapy. This highlights the important role of diagnostic stewardship in optimizing testing

algorithms and preventing unnecessary testing in cases where a microbiological

diagnosis has already been established. In this study, pre-approval practices varied

from one institution to the next with no center requiring ID pre-approval at the beginning

of the study. Policies changed over time with two centers (SHC and UCLA) requiring

pre-approval by microbiology or ID in the later phase of this study. In situations where

positive conventional test results precede the mNGS result, patient management (i.e.,

escalation or de-escalation of therapy) should be considered based on conventional

microbiological diagnostics rather than waiting for confirmatory mNGS result.

Microbiology laboratory or ID pre-approval of mNGS send-out requests represent an

important intervention component to reduce unnecessary testing in such cases.

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The finding that 6/82 (7.3%) of Karius test results in this study led to positive

clinical impact is in contrast to that recently reported in a single pediatric center in

Chicago where 56/100 (56.0%) test results were considered clinically actionable [10].

This discrepancy may have been driven by differences in patient selection, in addition to

differences in outcome assessment methodology. For instance, in our study, clinical

impact was solely based on the treating team’s interpretation of Karius results and their

subsequent management decisions; in contrast, in the above study, clinical adjudication

was performed where ‘clinical relevance’ was unclear from the medical record. Inherent

inter-provider interpretation differences of whether Karius results represent a clinically

significant pathogen or colonization also likely contributed to findings, including the fact

that not all results were reviewed by ID. Other differences between studies included

variability in testing indication, proportion of ordering infectious diseases providers, and

proportion of patients with a known microbiological diagnosis pre-Karius testing. The

low clinical impact proportion in the present study was closer to that reported for mNGS

on CSF where 8/204 (3.9%) of tests led to positive clinical impact [8].

This study was strengthened by its multicenter design including 5 adult and

pediatric sites, and by its assessment of real-world practice in presenting data from

provider-initiated testing. However, there are limitations. Firstly, although we made

significant effort to standardize the definition and application of clinical impact criteria,

some of the cases included were very complex medically and interpretation of clinical

impact may have been challenging to ascertain. The definition of impact was based on

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how providing physicians interpreted and reacted to test results. This may have over or

underestimated the real impact of the Karius test result. Standardized definitions of

impact across studies, in addition to more comprehensive and objective measures of

impact (for example, including antibiotic use, hospital length-of-stay, laboratory testing

utilization) are needed to accurately quantify and compare clinical impact. Secondly,

approval criteria for send-out testing varied by institution and time period such that

patient populations may have varied in their pre-test probability which may have

influenced the assessment of clinical impact. Finally, given this was a convenience

sample, not all patient populations are represented equally. For example, only one HIV-

1-positive individual was included, which limits generalizability to this patient population.

In conclusion, plasma mNGS represents an attractive diagnostic modality due to

its noninvasive nature and potential to provide an early and actionable diagnosis.

However, this study has shown that the real-world impact of the Karius test as currently

used in clinical practice is limited given that this testing resulted in negative impact or no

impact in addition to conventional results in the majority (92.7%) of patients. Further

studies are needed to better understand which patient populations may potentially

benefit, define the complementary role of mNGS to conventional microbiological

methods, and improve diagnostic interpretation algorithms. Furthermore, institutions

should adopt an active diagnostic stewardship role to ensure optimal use of mNGS.

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Funding

This work was supported in part by the Canadian Institutes for Health Research

201711HIV-398347-295229 to C.A.H..

Potential conflicts of interest. N.B. has received research funding from Karius from

2015-2016. No conflict for all other authors.

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References

1. Chiu CY, Miller SA. Clinical metagenomics. Nat Rev Genet 2019; 20(6): 341-55.

2. Blauwkamp TA, Thair S, Rosen MJ, et al. Analytical and clinical validation of a microbial

cell-free DNA sequencing test for infectious disease. Nat Microbiol 2019; 4(4): 663-74.

3. Hong DK, Blauwkamp TA, Kertesz M, Bercovici S, Truong C, Banaei N. Liquid biopsy for

infectious diseases: sequencing of cell-free plasma to detect pathogen DNA in patients

with invasive fungal disease. Diagn Microbiol Infect Dis 2018; 92(3): 210-3.

4. Armstrong AE, Rossoff J, Hollemon D, Hong DK, Muller WJ, Chaudhury S. Cell-free DNA

next-generation sequencing successfully detects infectious pathogens in pediatric

oncology and hematopoietic stem cell transplant patients at risk for invasive fungal

disease. Pediatr Blood Cancer 2019; 66(7): e27734.

5. Farnaes L, Wilke J, Ryan Loker K, et al. Community-acquired pneumonia in children: cell-

free plasma sequencing for diagnosis and management. Diagn Microbiol Infect Dis 2019;

94(2): 188-91.

6. Fung M, Zompi S, Seng H, et al. Plasma Cell-Free DNA Next-Generation Sequencing to

Diagnose and Monitor Infections in Allogeneic Hematopoietic Stem Cell Transplant

Patients. Open Forum Infect Dis 2018; 5(12): ofy301.

7. Zhou Y, Hemmige V, Dalai SC, Hong DK, Muldrew K, Mohajer MA. Utility of Whole-

Genome Next-Generation Sequencing of Plasma in Identifying Opportunistic Infections

in HIV/AIDS. The Open AIDS Journal 2019; 13(1): 7-11.

8. Wilson MR, Sample HA, Zorn KC, et al. Clinical Metagenomic Sequencing for Diagnosis of

Meningitis and Encephalitis. N Engl J Med 2019; 380(24): 2327-40.

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9. Miller S, Chiu C, Rodino KG, Miller MB. Should we be performing metagenomic next-

generation sequencing for infectious disease diagnosis in the clinical laboratory? J Clin

Microbiol 2019.

10. Rossoff J, Chaudhury S, Soneji M, et al. Noninvasive Diagnosis of Infection Using Plasma

Next-Generation Sequencing: A Single-Center Experience. Open Forum Infect Dis 2019;

6(8).

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Table 1. Standardized criteria to assess clinical impact of the Karius test per the treating team.

Category of

clinical impact

Change in

management Clinical impact type

Positive Yes

New diagnosis based on Karius result and not confirmed by conventional

microbiological methods

Earlier diagnosis based on Karius result and later confirmed by conventional

microbiological methods

Karius result enabled avoidance of invasive surgical biopsy

Karius result enabled initiation of appropriate therapy

Karius result enabled de-escalation of therapy

Karius result enabled escalation of therapy

No Karius result confirmed clinical diagnosis

Negative Yes

Karius result led to unnecessary treatment

Karius result led to additional unnecessary diagnostic investigations

Karius result led to longer length-of-stay

None No Karius result showed new organism but result not acted upon

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Karius result confirmed conventional microbiological diagnosis and not acted upon

Karius test result was negative and not acted upon

Patient died before Karius result available

Indeterminate

Yes

Could not determine clinical impact from chart review No

Indeterminate

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Table 2. Demographic and clinical characteristics of patients included in this study. CUMC SHC UCLA UT CHLA Total (%)

Number of patients 5 21 50 3 3 82

Number of tests 6 22 64 3 3 98

Mean Age (SD) 12.0 (5.8) 36.2 (24.3) 20.5 (20.0) 70.7 (8.4) 4 (0) 25.2 (23.3)

Male sex (%) 2 (40.0) 15 (71.4) 29 (58.0) 3 (100) 3 (100) 52 (63.4)

Immunosuppression (%) 5 (100) 14 (66.7) 30 (60.0) 3 (100) 1 (33.3) 53 (64.6)

Bone marrow transplant (%) 4 (80.0) 1 (4.8) 2 (4.0) 1 (33.3) 0 8 (9.8)

Solid organ transplant (%) 0 3 (14.3) 11 (22.0) 0 0 14 (17.1)

Active malignancy (%) 1 (20.0) 3 (14.3) 4 (8.0) 2 (66.7) 0 10 (12.2)

HIV-1 (%) 0 1 (4.8) 0 0 0 1 (1.2)

Primary immunodeficiency (%) 0 3 (14.3) 7 (14.0) 0 1 (33.3) 11 (13.4)

Other (%) 0 3 (14.3) 6 (12.0) 0 0 9 (11.0)

Mean turnaround time

in days (SD)

2.8 (0.8) 3.3 (1.9) 3.0 (0.9) 4.3 (0.6) 2.3 (0.6) 3.1 (1.2)

Test indication for first test

per patient (%)

- - - - - -

Diagnosis 5 (100) 19 (90.5) 46 (92.0) 3 (100) 3 (100) 76 (92.7)

Rule-out infection 0 1 (4.8) 0 0 0 1 (1.2)

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Monitoring 0 1 (4.8) 1 (2.0) 0 0 2 (2.4)

Unknown 0 0 3 (6.0) 0 0 3 (3.7)

Positive test results for first

test per patient (%)

5 (100) 12 (57.1) 28 (56.0) 3 (100) 2 (66.7) 50 (61.0)

CHLA: Children’s Hospital of Los Angeles; CUMC: Columbia University Medical Center; HIV-1: human immunodeficiency virus type 1; SD: Standard deviation; SHC: Stanford Health

Care; UCLA: University of California, Los Angeles; UT: University of Utah.

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Table 3. Clinical data for cases with clinical impact after Karius test result. Study

ID

Age/sex Comorbidities Clinical

syndrome

Karius test result

(molecules per

microliter)

Clinical

impact

per

treating

team

Clinical impact

type

Detail

SHC-

11

57M HIV/AIDS, CAD, CKD Suspected

mycotic

aneurysm

Enterococcus

faecalis

(344)

Prevotella

melaninogenica

(176)

Positive New diagnosis

and de-

escalation of

therapy

Karius test result was used to de-

escalate chronic suppressive

therapy to amoxicillin-clavulanic

acid, after a 6-week course of

empiric vancomycin and

ceftriaxone. Nine months later, after

multiple clinical recurrences,

vertebral tissue biopsy grew

Mycobacterium avium complex.

SHC-

13

47F s/p combined heart and

kidney transplant (2018)

for anthracycline-induced

cardiomyopathy, history

of ALL

Acute flaccid

paralysis (AFP)

Aspergillus

fumigatus

(qualitative result)

None/

Positive

None for AFP.

Earlier diagnosis

and initiation of

therapy for

fungal infection

Karius test identified Aspergillus

fumigatus earlier than conventional

methods and prompted chest

imaging and respiratory cultures

that confirmed invasive pulmonary

aspergillosis. Serum

galactomannan had been elevated

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(>2.00) in the week prior to Karius

test.

UCLA-

3

2M Idiopathic acute liver

failure, aplastic anemia,

HLH

Recurrent fever Candida tropicalis

(6,670)

Positive Earlier diagnosis

and initiation of

therapy

Karius test result was used to

initiate antifungal therapy with

caspofungin 5 days earlier than the

time of diagnosis by blood culture.

UCLA-

20

10F Liver/small

bowel/colon/pancreas

transplant (2018), TPN-

dependent short gut

syndrome

Jejunal

perforation with

necrotic fascia,

subcutaneous

abscess,

perinephric fluid

collection

Rhizopus oryzae

(169,935)

Stenotrophomonas

maltophilia

(2,649)

E. coli

(234)


not made by

conventional

methods and

escalation of

therapy

Karius test diagnosed invasive

mucormycosis, which was not

made through conventional

methods and which enabled

targeted antifungal therapy. The

patient was not treated for the

bacterial organisms detected.

UCLA-

47

6wk-old

M

Newborn by emergency

caesarean section,

TAPVR s/p repair

Multi-organ

dysfunction

syndrome,

coagulopathy,

respiratory failure

Negative Positive De-escalation of

therapy

Karius test result was negative and

supported the clinical suspicion that

patient's clinical decompensation

was not infectious in etiology. This

prompted discontinuation of empiric

cefepime.

UT-

2

75M Metastatic carcinoma of

unknown primary

Multifocal

abscesses (lung,

liver and kidney)

Streptococcus

parasanguinis

(372)


not made by

conventional

Karius test identified mixed oral

flora, which was consistent with a

periodontal source of infection. This

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Morococcus

cerebrosus

(212)

Neisseria sicca

(96)

Veillonella parvula

(64)

Prevotella

melaninogenica

(36)

Propionibacterium

acidifaciens

(24)

EBV

(23)

methods and

de-escalation of

therapy

allowed narrowing of antibacterial

therapy from

vancomycin/piperacillin-tazobactam

to amoxicillin-clavulanic acid for a

total duration of 6 weeks of therapy.

CUMC-

5

17F GATA2-MDS s/p PBSC

transplant (2018)

Fever, buccal

lesions, hepatic

lesions

Trichosporon asahii

(1,786)

Enterococcus

faecium

(1,567)

Mucor indicus

(1,199)

Negative Unnecessary

treatment

Karius test result prompted

additional antibacterial coverage

that was not thought to be

warranted per ID team. Patient was

already known to have invasive

mucormycosis.

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CMV

(763)

Rhizopus delemar

(107)

Candida glabrata

(75)

Veillonella parvula

(47)

SHC-

8

39F CVID Chronic low-

grade fever

Prevotella bivia

(23)

Streptococcus

pseudopneumoniae

(23)

Bacteroides fragilis

(21)

Negative Additional

diagnostic

investigations

Karius test result prompted an ED

visit, and subsequent outpatient

consultation with ID to outline

management. The patient was not

treated for the Karius test result.

SHC-

14

20M Refractory and recurrent

marginal cell MALT-like

lymphoma of ileum and

colon

Investigation of

lymphoma

etiology

Trichoderma

atroviride

(695)

Enterobacter

cloacae complex

(96)

Negative Additional

diagnostic

investigations

Karius test result prompted concern

by the treating team about a false

positive result. Targeted fungal

sequencing on plasma was

performed to try to disprove result

and was negative. The patient was

not treated for the Karius test result.

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AIDS: acquired immune deficiency syndrome; ALL: acute lymphoblastic leukemia; AML: acute myeloid leukemia; AFP: acute flaccid paralysis; CAD: coronary artery disease; CHLA:

Children’s Hospital of Los Angeles; CKD: chronic kidney disease; CUMC: Columbia University Medical Center; CVID: common variable immune deficiency; ED: emergency

department; F: female; GATA-2: GATA-2 gene; HIV: human immunodeficiency virus; HLH: hemophagocytic lymphohistiocytosis; ID: infectious diseases; M: male; MALT: mucosa-

associated lymphoid tissue; MDS: myelodysplastic syndrome; PBSC: peripheral blood stem cell; SHC: Stanford Health Care; s/p: status post; TAPVR: total anomalous pulmonary

venous return; TPN: total parenteral nutrition; UCLA: University of California, Los Angeles; UT: University of Utah; wk: week

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Documents

Department of Pathology, Stanford University School of ......DNA and RNA in respiratory secretions (IDbyDNA, San Francisco, CA) [1]. Of these, the Karius test has been commercially